Generation of Biomarkers for the Detection of ADP-ribosylated Proteins During Cellular Stress

ADP-ribosylation of proteins is a reversible post-translational modification in which the ADP-ribose (ADPr) moiety of NAD+ is transferred to a specific amino acid of the acceptor protein. Up to now, more than 200 proteins, mostly localized in the nucleus, were identified to be mono- or poly-ADP- ribosylated. However, a comprehensive understanding of the molecular mechanism of the ADP-ribosyltransferase reaction and of the cellular functions of ADP-ribosylation is still missing.

The aim of this project is to establish fast and reliable assays for the identification and quantification of specific ADP-ribosylated proteins in physiological and pathological conditions by using an MS/MS approach. We will first characterize proteins that are ADP-ribosylated upon oxidative stress. Bioinformatic analyses and assay optimizations will then identify marker peptides that will be quantified by LC-SRM MS/MS in cell lysates (no treatment vs. oxidative stress conditions). Finally, the biomarkers will be validated using various cell lines exposed to different stress conditions.

In the future, the newly developed methods for the quantification of specific ADP-ribose modifications of proteins can potentially be applied for the stratification and diagnosis of pathological conditions. Additionally, it will help to further understand the role of this PTM in the cell.